of response occurs only if the anti-I-A k reagent is directed against determinants ... unique and essential role of accessory cell Ia determinants that must be recognized by. T helper cells for ... over Sephadex G-10 columns, and then by depleting the G-10-passed spleen cells oft ceils by ... All points shown in each experiment.
MAJOR
HISTOCOMPATIBILITY
RESTRICTED
COMPLEX-
SELF RECOGNITION
A Monoclonal Anti-I-A k Reagent Blocks H e l p e r T Cell R e c o g n i t i o n o f Self M a j o r H i s t o c o m p a t i b i l i t y C o m p l e x D e t e r m i n a n t s By RICHARD J. HODES, KAREN S. HATHCOCK, AND ALFRED SINGER From the Immunology Branch, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20205
Major histocompatibility complex (MHC) 1 genes, in particular those encoded in the I region of the murine H-2, have been shown to be of functional importance in several aspects of immune response regulation. Thus, I region-encoded immune response (It) genes determine responsiveness or unresponsiveness to a number of antigens (1); and T cell recognition of I region gene products is critical for the H - 2 restricted cell interactions that mediate a number of immune responses (2-6). Although these I region gene functions are well established, the precise identification of the gene products mediating these functions remains uncertain. In contrast, there exists a class of I region gene products, the Ia antigens, for which extensive serologic and biochemical characterization has been accomplished (7-9). The functional role of Ia antigens in immune responses remains undetermined, however, and has been a subject of wide speculation. The present report represents an attempt to assess the functional significance of Ia antigens in the regulation of antibody responses studied in vitro. A system of primary in vitro antibody responses has been characterized in which responses to soluble trinitrophenyl (TNP)-protein conjugates require the cooperation of T cells, B cells, and accessory cells (6). In these responses, a strict requirement has previously been demonstrated for helper T cell recognition of K or L A encoded determinants expressed on accessory cells (6, 10). In the present studies, the functional role of serologically defined cell surface Ia antigens has been investigated. Using Iaspecific serologic reagents as probes, the present study demonstrates that: (a) a monoclonal anti-Ia reagent specific for a product of I-A k is capable of inhibiting antibody responses in vitro; and that (b) such inhibition occurs at the level of an I-A k product expressed by accessory cells. Moreover, (c) inhibition by this anti-Ia reagent occurs only for responses in which the I-A product on accessory cells is actively recognized by helper T cells as self; and (d) for Ir gene controlled responses, inhibition of response occurs only if the anti-I-A k reagent is directed against determinants 1Abbreviations used in this paper: B10, C57BL/10; C', complement; FI, (B10 x B10.A)FI; FCS, fetal calf serum; Ia, 1 region associated; b, immune response; KLH, keyhole limpet hemocyanin; MEM, Eagle's minimal essential medium; MHC, major histocompatibilitycomplex; PFC, plaque-formingcell; RAMB, rabbit anti-mouse brain serum; SAC, spleen adherent cell; SRBC, sheep erythrocyte;TNP, trinitrophenyl.
THE JOURNAL OF EXPERIMENTAL MEDICINE • VOLUME 152, 1980
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ANTI-Ia INHIBITION OF T CELL RECOGNITION
encoded by the responder haplotype. These findings are interpreted as evidence for a u n i q u e a n d essential role of accessory cell Ia d e t e r m i n a n t s that must be recognized by T helper cells for the generation of a n t i b o d y responses. Materials and Methods Animals. C57BL/10 (BI0), BI0.A, and (B10 × BI0.A)F1 (F1) mice were obtained from The Jackson Laboratory, Bar Harbor, Maine. B10.A(4R), B10.A(5R), and BI0.MBR mice were generously provided from the colony of Dr. David Sachs (National Institutes of Health, Bethesda, Md.). Adult males 2-5 mo old were used in all experiments. Chimeras. Recipient mice were irradiated with 950R X-ray and reconstituted 2-6 h later with 15 × 106 bone marrow cells that had been pretreated with rabbit anti-mouse brain serum (RAMB) and complement (C'). The RAMB reagent employed has been extensively characterized and is cytotoxic for T cells (11) with no detectable anti-stem cell activity (6). Chimeras are designated as bone marrow donor ~ irradiated recipient. Spleen cells were obtained from each chimera no earlier than 2 mo postirradiation, and were individually typed by indirect immunofluorescence using H-2 specific reagents as previously described (6). By such testing, spleen cells from each chimera employed were of donor origin without detectable (
